Hb Rancho Mirage was detected in a 17-year-old male in association with a mild anemia. Hemoglobin electrophoresis revealed the variant had a mobility between Hbs A and J on cellulose acetate (pH 8.6) and a mobility like Hb F on citrate agar (pH 6.4). A substitution of His----Asp was found at position 143 in the beta chain, a residue that contributes to the anionic 2,3-DPG binding site in Hb. This variant exhibited normal oxygen affinity at physiologic pH and reduced affinity at alkaline pH. This suggested a subtle shift in the allosteric equilibrium due most likely to the introduction of a negative charge that stabilized the 2,3-DPG pocket. Both homotrophic (heme-heme) and heterotropic (2,3-DPG and protons) effects were reduced; this might be a consequence of an alteration in the carboxyl terminal region of the beta-subunits. Although a His----Asp substitution would be considered to cause reasonable disruption of the 2,3-DPG and C-terminal conformation of the beta- subunits, the properties of Hb Rancho Mirage suggest that, in fact, there appear to be no major perturbation of the critical C-terminal residues.